Biotechnology of α-linolenic acid in oilseed rape(Brassica napus)using FAD2 and FAD3 from chia(Salvia hispanica)

α-Linolenic acid (ALA,18:3Δ9,12,15) is an essential fatty acid for humans since it is the precursor for the biosynthesis of omega-3 long-chain polyunsaturated fatty acids (LC-PUFA).Modern people generally suffer from deficiency of ALA because most staple food oils are low or lack ALA content.Biotechnological enrichment of ALA in staple oil crops is a promising strategy.Chia (Salvia hispanica) has the highest ALA content in its seed oil among known oil crops.In this study,the FAD2 and FAD3 genes from chia were engineered into a staple oil crop,oilseed rape (Brassica napus),via Agrobaterium tumefaciens-mediated transformation of their LP4-2A fusion gene construct driven by the seed-specific promoter PNap A.In seeds of T0,T1,and T2 lines,the average ALA contents were 20.86,23.54,and 24.92%,respectively,which were 2.21,2.68,and 3.03 folds of the non-transformed controls (9.42,8.78,and 8.22%),respectively.The highest seed ALA levels of T0,T1,and T2 plants were 38.41,35.98,and 39.19%respectively,which were 4.10–4.77 folds of the respective controls.FA-pathway enzyme genes (Bn ACCD,Bn FATA,Bn SAD,Bn SCD,Bn DGAT1,Bn DGAT2,and Bn DGAT3) and positive regulatory genes (Bn WRI1,Bn LEC1,Bn L1L,Bn LEC2,Bn ABI3,Bnb ZIP67,and Bn MYB96) were all significantly up-regulated.In contrast,Bn TT1,Bn TT2,Bn TT8,Bn TT16,Bn TTG1,and Bn TTG2,encoding negative oil accumulation regulators but positive secondary metabolism regulators,were all significantly down-regulated.This means the foreign Sh FAD2-Sh FAD3 fusion gene,directly and indirectly,remodeled both positive and negative loci of the whole FA-related network in transgenic B.napus seeds.